Open AccessThree dimensional single molecule localization using a phase retrieved pupil function
Author(s) -
Sheng Liu,
Emil B. Kromann,
Wesley D. Krueger,
Joerg Bewersdorf,
Keith A. Lidke
Publication year - 2013
Publication title -
optics express
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.394
H-Index - 271
ISSN - 1094-4087
DOI - 10.1364/oe.21.029462
Subject(s) - point spread function , optics , pupil , phase retrieval , optical transfer function , cardinal point , microscope , pupil function , computer science , phase (matter) , spatial frequency , image resolution , physics , fourier transform , quantum mechanics
Localization-based superresolution imaging is dependent on finding the positions of individual fluorophores in a sample by fitting the observed single-molecule intensity pattern to the microscope point spread function (PSF). For three-dimensional imaging, system-specific aberrations of the optical system can lead to inaccurate localizations when the PSF model does not account for these aberrations. Here we describe the use of phase-retrieved pupil functions to generate a more accurate PSF and therefore more accurate 3D localizations. The complex-valued pupil function contains information about the system-specific aberrations and can thus be used to generate the PSF for arbitrary defocus. Further, it can be modified to include depth dependent aberrations. We describe the phase retrieval process, the method for including depth dependent aberrations, and a fast fitting algorithm using graphics processing units. The superior localization accuracy of the pupil function generated PSF is demonstrated with dual focal plane 3D superresolution imaging of biological structures.